We are studying a newly-identified F-box receptor for ubiquitin- mediated protein degradation, Partner of paired (Ppa), whose properties suggest roles in nuclear cycle regulation as well as in segmental patterning of gene expression during Drosophila embryogenesis. Preliminary studies suggest that ppa promotes progression through mitosis (see Aim 1), and we hypothesize that this occurs through regulation of cyclin degradation (Aim 2). We have established previously that Ppa coordinates degradation of the Pax transcription factor, Paired. We hypothesize that after the first 13 rapid nuclear cycles, localized elimination of ppa expression during the long interphase of cycle 14 creates a developmental window in which Prd can function in the absence of mitosis. We hypothesize that this is important because Prd would otherwise be detrimental to mitosis (Aim 3). Our specific aims are: (1) To establish whether ppa promotes mitosis. We will test whether localized ppa expression overlaps in time and space the orchestrated domains of mitosis 14. We will examine whether in ppa mutants, progression of mitosis 14 in these domains is compromised. We will also build on preliminary results which suggest that progression through mitosis 13 is delayed in ppa mutants. (2) To test whether Ppa regulates Drosophila cyclins. We will determine whether Ppa interacts with particular Drosophila cyclins by testing for protein interactions in two-hybrid and co-immunoprecipitation experiments, and by testing for genetic interactions in gene dosage experiments in which ppa and cyclin levels are altered. Examination of cyclin levels by western analysis in ppa mutants and in embryos with ppa over-expression transgenes will test whether ppa can regulate cyclin protein levels. (3) To test whether Prd function is detrimental to mitosis. By comparing the spatial expression of Prd and Ppa with the localized mitotic domains normally observed in nuclear cycle 14, we will test whether Prd function and mitosis are mutually exclusive, as suggested by preliminary experiments. We will then test, using prd over-expression transgenes, whether ectopic Prd function modulates nuclear cycling. Our proposed analysis will provide insights into the multi-layered regulatory network that coordinates nuclear cycling and pattern formation in Drosophila development, and will suggest possible regulatory controls for future analysis of vertebrates.